The aerosol performance, physical properties and formation process of two corticosteroids (beclomethasone dipropionate and fluticasone propionate) and caffeine (active pharmaceutical ingredients: APIs) from ethanol-based pressurized metered dose inhaler solution formulations, containing various ethanol fractions, were evaluated using cascade impaction, thermal analysis and scanning electron microscopy. In general, the final aerosol particle size distribution (post USP induction port) was unaffected by ethanol concentration (mass median aerodynamic diameter and geometric standard deviation values for each formulation were independent of ethanol % (w/w) in the initial formulation). However, ethanol concentration directly affected the percentage of particles that passed the USP induction, resulting in a significant decrease in fine particle fraction, across all formulations, as ethanol was increased. Thus it can be concluded that particle size is governed by initial droplet diameter and API concentration, while performance is governed by drying time. The physico-chemical properties and morphology of the dried API particles, collected from cascade impactor stages, showed that the solid state was related to the glass transition temperature (Tg) and, to some extent, the saturated hydrofluoroalkane propellant (HFA)/ethanol solubility of the APIs. The low Tg API caffeine, with high HFA solubility resulted in crystalline particles, while the high Tg corticosteroids were amorphous. Furthermore, the final structure of the particles was dependent on the ethanol concentration and drying kinetics after initial droplet formation. This study has shown that the solid-state physico-chemical properties and morphology of particles is intrinsically linked to the API properties and drying kinetics of the propellant/co-solvent. These variations in aerosol efficiency, particle morphology and solid-state characteristics may have direct effects on drug efficacy and bioavailability after deposition in the lung.

Purpose: To investigate the influence of different actuator materials and nozzle designs on the electrostatic charge properties of a series of solution metered dose inhaler (pMDI) aerosols.

Methods: Actuators are manufactured with two different nozzle designs (flat and cone) using five different materials selected from the triboelectric series (Nylon, Polyethylene terephthalate, Polyethylene – High density, Polypropylene copolymer and Polytetrafluoroethylene). The electrostatic charge profiles of pMDI formulations containing beclomethasone dipropionate (BDP) as model drug, in HFA-134a propellant, with different concentrations of ethanol were studied with different actuator designs. Electrostatic measurements were taken using a modified electrical low-pressure impactor (ELPI) and the deposited drug mass was assayed chemically using HPLC.

Results: The charge profiles of HFA 134a propellant alone have shown strong electronegativity with all actuator materials and nozzle designs, at an average of –1531.34 pC ± 377.34. The presence of co-solvent ethanol significantly reduced the negative charge magnitude of HFA 134a, with 15% ethanol producing a net charge close to neutral for all actuator designs. BDP reduces the suppressing effect of ethanol on the negative charging property of the propellant. For all tested formulations, the flat nozzle design shown no significant differences between different actuator materials, where for the cone designs the net charge profile ranking follows the triboelectric series.  

Conclusion: The electrostatic charging profiles arising from a solution pMDI formulation containing BDP and ethanol as co-solvent can be significantly influenced by the actuator material, nozzle design and formulation components. Ethanol concentration appears to have the most significant impact. Furthermore, it has been shown that BDP interactions with ethanol and HFA could have an influence on the electrostatic charge of the aerosol. By choosing a different combination of actuator materials and orifice design, the fine particle fractions of the formulation can be altered.

Objectives: The formulation of a clarithromycin (CLA) pressurised metered dose inhalers (pMDIs) solution formulation opens up exciting therapeutic opportunities for the treatment of inflammation in chronic obstructive lung diseases. In this study we have formulated and tested a low dose macrolide formulation of CLA for treatment of inflammation and studied its physico-chemical and aerosol properties.

Methods: The system was characterised for in vitro aerosol performance using an Anderson cascade impactor. Short-term chemical and physical stability was assessed by dose content uniformity over a range of temperatures. Standard physico-chemical characteristics were also investigated using scanning electron microscopy, thermo analysis and laser diffraction techniques. 

Results and discussion: The formulation had a relatively high fine particle fraction (47%) and produced a particle size distribution suitable for inhalation drug delivery. Particles had an irregular morphology and were predominately amorphous. Furthermore, the short-term stability showed the formulation to be stable from 4-37°C.

Conclusions: This study demonstrated the feasibility of formulating a solution-based pMDI containing of CLA for the treatment of lung inflammatory diseases.

Purpose: The potential for rifapentine-containing oral therapeutic regimens to significantly shorten the current six-month anti-tubercular treatment regimen is confounded by high plasma protein binding of rifapentine. Inhaled aerosol delivery of rifapentine, a more potent anti-tubercular antibiotic drug, in combination with other first- line antibiotics may overcome this limitation to deliver a high drug dose at the pulmonary site of infection. Method: A formulation consisting of rifapentine, moxifloxican and pyrazinamide with and without leucine was prepared by spray-drying. This formulation was assessed for its physico-chemical properties, and in vitro aerosol performance and antimicrobial activity. Results: The antibiotic powders, with and without leucine, had similar median aerodynamic diameters of 2.58 ± 0.08 μm and 2.51 ± 0.06 μm, with a relatively high fine particle fraction of 55.5 ± 1.9% and 63.6 ± 2.0%, respectively. Although the powders were mostly amorphous, some crystalline peaks associated with the δ polymorph for the spray-dried crystalline pyrazinamide were identified. Conclusions: Stabilisation of the powder with 10% w/w leucine and protection from moisture ingress was found to be necessary to prevent the crystallisation of pyrazinamide after long-term storage. In vitro biological assays indicated antimicrobial activity was retained after spray-drying. Murine pharmacokinetic studies are currently underway.

A critical problem associated with poor water-soluble drugs is their low and variable bioavailability, which is derived from the slow dissolution and erratic absorption. Nano-formulation has been identified as one approach to enhance the rate and extent of drug absorption for compounds that demonstrate limited water solubility. This study aimed to investigate the physico-chemical variables that affect the manufacture, dissolution and consequent bioavailability of wet-milled clarithromycin (CLA) nanoparticles, a macrolide antibiotic. CLA nanoparticles were prepared using wet milling method followed by freeze-drying. Different stabilizer systems, consisting of surfactants and polymers alone or their combinations were studied to determine the optimum conditions for producing nano-sized CLA particles. In vitro characterizations of the CLA nanoparticles were performed using photon correlation spectroscopy, X-ray powder diffraction, differential scanning calorimetry and dissolution efficiency test. Results showed that in general the wet milling process did not modify the crystallinty of the CLA nanoparticles. The poloxamers and poly vinyl alcohol (PVA) stabilizers resulted in nanoparticles with the smallest particle size and best dissolution rates. Furthermore, poloxamers F68 and F127, and PVA stabilizers demonstrated the best performance in increasing dissolution efficacy.

The aim of the present study was to develop controlled release inhalable lipid microparticles (LMs) loaded with the antioxidant flavonoid, quercetin and to investigate the interaction of these microparticles with A549 pulmonary alveolar epithelial cells. The LMs were produced using different lipidic materials and surfactants, by melt emulsification followed by a sonication step. The most efficient modulation of the in vitro release of quercetin was achieved when LMs were prepared with tristearin and hydrogenated phosphatidylcholine; which were used for subsequent studies.

These LMs exhibited a quercetin loading of 11.8±0.3%, and a volume median diameter, determined by laser diffraction, of 4.1±0.2 μm.  Moreover, their mass median aerodynamic diameter (4.82 ±0.15 μm) and fine particle fraction (27.2±3.9%), as measured by multi-stage liquid impinger, were suitable for pulmonary delivery. Quercetin was found to be highly unstable (complete decomposition within 6-h incubation) in Ham’s F-12 medium used for A549 cell culture. Degradation was markedly reduced (16.4% of the initial quercetin content still present after 24-h incubation) after encapsulation in the lipid system. 

Viability studies performed by lactate dehydrogenase assay, demonstrated that quercetin LMs showed no significant cytotoxicity on the A549 cells, over the concentration 0.1-5 μM. The uptake of quercetin by the A549 lung alveolar cells was also investigated.  After 4-h incubation, the accumulation of quercetin in the A549 cells was significantly higher (2.3-fold increase) for the microparticle entrapped flavonoid when compare to non-encapsulated quercetin. The enhanced intracellular delivery of quercetin achieved by the LMs is likely due to the flavonoid stabilization after encapsulation. 

Purpose: A new approach to delivering high doses of dry powder medicaments to the lung is presented. The Orbital dry powder device (DPI) is designed to deliver high-doses of drugs to the respiratory tract in a single dosing unit, via multiple inhalation manoeuvrers, overcoming the need to prime or insert multiple capsules.

Methods: The Orbital was tested in its prototype configuration and compared to a conventional RS01 capsule device. Three formulations were evaluated: 200 mg spray-dried ciprofloxacin formulation for respiratory infection, 200 mg spray-dried mannitol formulation for mucus clearance, and 100, 200 and 400 mg co-spray dried 1:8 formulations containing ciprofloxacin and mannitol as combination therapy. The systems were evaluated in terms of physico-chemical properties and tested using a multistage liquid impinger at 60 L/min. Emptying rates were evaluated and the aerosolisation performance compared to 10 capsules used sequentially in the RS01. 

Results and Discussion: The systems were different in terms of morphology, thermal response, moisture sorption and stability; however, they had similar sizes when measured by laser diffraction making them suitable for comparison in the Orbital and RS01 devices. The aerosolisation performance from the Orbital and RS01 was dependent upon the formulation type; however, the fine particle fraction (FPF) produced by the Orbital device was higher than the RS01.  The FPF for ciprofloxacin, mannitol and co-spray dried formulation were 67.1±1.8, 47.1 ±2.2 and 42.0 ±1.8, respectively. For the Orbital, 90% of the loaded dose was delivered within 10 inhalation maneuvers, with the profile being dependent upon the formulation type. 

Conclusion: The Orbital provides a means of delivering high doses of medicine to the respiratory tract through multiple breath maneuvers after a single actuation. This approach will allow the delivery of a wide range of high-pay load formulations (>100 mg) for the treatment of a variety of lung disorders. To date no such passive device exists that meet these crucial criteria.

Inhalation of antibiotics and mucolytics are the most important combination of inhaled drugs for chronic obstructive lung diseases and have become a standard part of treatment. However, it is yet to be determined whether the administration of a mucolytic has an effect on the transport rate of antibiotics across the airway epithelial cells. Consequently, the aim of this study was to investigate the effects of inhalation dry powder, specifically mannitol, on ciprofloxacin transport using Calu-3 air interface cell model. Transport studies of ciprofloxacin HCl were performed using different configurations including single spray-dried ciprofloxacin alone, co-spray dried ciprofloxacin with mannitol and deposition of mannitol prior to ciprofloxacin deposition. To understand the mechanism of transport and interactions between the drugs, pH measurements of apical surface liquid and further transport studies were performed with ciprofloxacin base, with and without the presence of ion channel/transport inhibitors such as disodium cromoglycate and furosemide. Mannitol was found to delay absorption of ciprofloxacin HCl through the increase in ASL volume and subsequent reduction in pH. Conversely, ciprofloxacin base had a higher transport rate after mannitol deposition. This study clearly demonstrates that the deposition of mannitol prior to ciprofloxacin on the air-interface Calu-3 cell model has an effect on its transport rate. This was also dependent on the salt form of the drug and the timing and sequence of formulations administered.

Liposomal ciprofloxacin formulations have been developed with the aim of enhancing lung residence time, thereby reducing the burden of inhaled antimicrobial therapy which requires multiple daily due to rapid absorptive clearance of antibiotics from the lungs. However, there is a lack of a predictive methodology available to assess controlled release inhalation delivery systems and their effect on drug disposition. In this study three ciprofloxacin formulations were evaluated: a liposomal formulation, a solution formulation and a 1:1 combination of the two (mixture formulation). Different methodologies were utilised to study the release profiles of ciprofloxacin from these formulations: (i) membrane diffusion, (ii) air-interface Calu-3 cells, and (iii) isolated perfused rat lungs. The data from these models were compared to the performance of the formulations in vivo. The solution formulation provided the highest rate of absorptive transport followed by the mixture formulation, with the liposomal formulation providing substantially slower drug release. The rank- order of drug release/transport from the different formulations was consistent across the in vitro and ex-vivo methods, and this was predictive of the profiles in vivo. The use of complimentary in vitro and ex-vivo methodologies provided a robust analysis of formulation behavior, including mechanistic insights, and predicted in vivo pharmacokinetics.

Introduction: The Calu-3 lung cell line has been shown to be a promising in vitro model of airway epithelia due to its similarity to in vivo physiology. Hence, over the last decade, it has found increasing applications in the pharmaceutical industry. 

Areas covered: This review focuses on the pharmaceutical applications of the Calu-3 cell line in areas such as drug transport, metabolism, controlled release studies, identification of possible drug-drug interactions, investigations into the mechanisms of action of active compounds and understanding of disease pathophysiology. The main findings of various studies, as well as the predictive potential of this model, are presented and discussed in this review.

Expert opinion: There is still a lack of mechanistic knowledge regarding transport of inhaled therapeutics across the lungs. Cell culture models such as Calu-3 provides a simple and reproducible system to study the underlying mechanisms by which inhaled therapeutics interact with the lungs. However, more complex systems that integrate particle deposition onto different cell culture systems may be useful in addressing some fundamental questions to generate a better understanding of determinants that influences pulmonary drug dissolution, absorption, metabolism and efficacy. Ultimately the use of the Calu-3 cell line provides a basic research tool that enables the development of safer and more effective inhaled therapeutics.

Australian data suggest up to 15% of people with intellectual disability (ID) have asthma. The inhaled route of administration is optimal for the management of obstructive airways diseases; however, correct inhaler use requires dexterity and particular breathing patterns and potentially represents a problem in this population due to physical and cognitive deficits. Understanding the nature and extent of inhaler use in persons with ID is important, as correct inhaler technique is imperative for optimal clinical outcomes; however, currently no evidence base exists to inform health professionals. This study describes respiratory medication use, reported prevalence of asthma, and asthma management practices undertaken in a community sample of Australian adults with ID. Results showed a prevalence of retrospectively reported asthma of 6%, with 86% of asthma patients prescribed inhaled medication. A review of patient records also indicated omission of some recommended asthma management strategies.

Background and objective: We aimed to assess whether co-deposition of a long-acting β2-agonist and a steroid affects their respective transport rates across epithelial cells.

Methods: Drug particles were deposited on the air-interface culture of Calu-3 cells using a twin stage impinger. We compared the transport rate of salmeterol and fluticasone from commercially available formulations (Serevent®, Flixotide® and Seretide®) across the epithelial cells. The transepithelial resistance of Calu-3 cells was measured before and after each deposition to monitor epithelial resistance.

Results: The co-deposition of salmeterol and fluticasone had no significant effect on transport of salmeterol through the cell layer, suggesting that salmeterol particles are deposited evenly across the cell line and each particle could have its own dissolution and uptake profile. In contrast, the rate of FP transport in presence of S was significantly lower (0.53 ± 0.20%) compared to the single FP formulation (2.36 ± 0.97%). Furthermore, the resistance of the epithelial cells was significantly increased after salmeterol deposition from both single and combination product.

Conclusions: Our data demonstrates that salmeterol may decrease the permeability of epithetical cells, resulting in slower fluticasone transport across Calu-3 epithelial monolayer. The subsequent increased residence time of fluticasone in the airways could prolong it’s anti-inflammatory effects.

Purpose: The aim of the present work was to develop solid lipid micro particles (SLMs), as dry powders containing quercetin for direct administration to the lung.

Methods: Quercetin microparticles were prepared by o/w emulsification via a phase inversion technique, using tristearin as the lipid component and phosphatidylcholine as an emulsifier. The quercetin SLMs were characterised for morphology, drug loading (15.5 % ± 0.6, which corresponded to an encapsulation efficiency of 71.4%.), particle size distribution, response to humidity, crystallinity, thermal behaviour and in vitro respirable fraction. Furthermore, the toxicity and the in vitro transport of the SLMs on an air liquid interface model of the Calu-3 cell line were also investigated using a modified twin-stage impinger apparatus.

Results: Results showed that quercetin SLMs could be formulated as dry powder suitable for inhalation drug delivery (20.5 ± 3.3% fine particle fraction ≤ 4.46 mm) that was adsorbed, via a linear kinetic model across the Calu-3 monolayer (22.32 ± 1.51 % over 4 hours). In addition, quercetin SLMs were shown to be non-toxic at the concentrations investigated. Interestingly, no apical to basolateral transport of the micronized quercetin was observed over the period of study.

Conclusions: These observations suggest quercetin diffusion was enhanced by the presence of the lipid/emulsifying excipients in the SLMs; however further studies are necessary to elucidate the exact mechanisms.

Ciprofloxacin is a well-established broad-spectrum fluroquinolone antibiotic that penetrates well into the lung tissues; still the mechanisms of its transepithelial transport are unknown. Contributions of specific transporters including multidrug efflux transporters, organic cation transporters and organic anion transporting polypeptide transporters on the uptake of ciprofloxacin were investigated in vitro using an air interface bronchial epithelial model. Our results demonstrate that ciprofloxacin is subject to predominantly active influx and a slight efflux component.

Two solution-based pressurised metered dose inhaler (pMDI) formulations were prepared such that they delivered aerosols with identical mass median aerodynamic diameters, but contained either beclomethasone dipropionate (BDP) alone (glycerol-free formulation) or BDP and glycerol in a 1:1 mass ratio (glycerol-containing formulation). The two formulations were deposited onto Calu-3 respiratory epithelial cell layers cultured at an air-interface. Equivalent drug mass (~1000 ng or ~2000 ng of the formulation) or equivalent particle number (1000 ng of BDP in the glycerol-containing vs. 2000 ng of BDP in the glycerol-free formulation) were deposited as aerosolised particles on the air interfaced surface of the cell layers. The transfer rate of BDP across the cell layer after deposition of the glycerol-free particles was proportional to the mass deposited. In comparison the transfer of BDP from the glycerol-containing formulation was independent of the mass deposited, suggesting that the release of BDP is modified in the presence of glycerol. The rate of BDP transfer (and the extent of metabolism) over 2 h was faster when delivered in glycerol-free particles, 465.01 ng ± 95.12 ng of the total drug (20.99 ± 4.29 %; BDP plus active metabolite) transported across the cell layer, compared to 116.17 ng ± 3.07 ng (6.07 ± 0.16 %) when the equivalent mass of BDP was deposited in glycerol-containing particles. These observations suggest that the presence of glycerol in the maturated aerosol particles may influence the disposition of BDP in the lungs.

A series of semi-empirical equations were utilised to design two solution based pressurised metered dose inhaler (pMDI) formulations, with equivalent aerosol performance but different physicochemical properties. Both inhaler formulations contained the drug, beclomethasone dipropionate (BDP), a volatile mixture of ethanol co-solvent and propellant (hydrofluoroalkane-HFA). However, one formulation was designed such that the emitted aerosol particles contained BDP and glycerol, a common inhalation particle modifying excipient, in a 1:1 mass ratio. By modifying the formulation parameters, including actuator orifice, HFA and metering volumes, it was possible to produce two formulations (glycerol-free and glycerol-containing) which had identical mass median aerodynamic diameters (2.4 mm  ± 0.1 and 2.5 mm  ± 0.2), fine particle dose (≤ 5 mm; 66 mg  ± 6 and 68 mg  ± 2) and fine particle fractions (28 % ± 2 and 30 % ± 1), respectively. These observations demonstrate that it is possible to engineer formulations that generate aerosol particles with very different compositions to have similar emitted dose and in vitro deposition profiles, thus making them equivalent in terms of aerosol performance. Analysis of the physicochemical properties of each formulation identified significant differences in terms of morphology, thermal properties and drug dissolution of emitted particles. The particles produced from both formulations were amorphous; however the formulation containing glycerol generated particles with a porous structure, while the glycerol-free formulation generated particles with a primarily spherical morphology. Furthermore, the glycerol-containing particles had a significantly lower dissolution rate (7.8% ± 2.1%, over 180 min) compared to the glycerol-free particles (58.0% ± 2.9%, over 60 min) when measured using a Franz diffusion cell.  It is hypothesised that the presence of glycerol in the emitted aerosol particles altered solubility and drug transport, which may have implications for BDP pharmacokinetics after deposition in the respiratory tract.

Silver nanoparticles (AgNPs) with size ranging from 7 to 70 nm were synthesized using ascorbic acid-citrate seed-mediated growth approach at room temperature. The 8 nm silver particles were prepared using gallic acid in alkaline conditions and used as seed to prepare AgNPs. The presence of ascorbic acid and citrate allows the regulation of size and size distribution of the nanoparticles. The increase in free silver ion-to-seed ratio (Ag+/Ag0) resulted in the changes of particle shape from spherical to pseudo-spherical and minor cylindrical shape. Further repetitive seeding approach resulted in the formation of pseudo-spherical particles with higher polydispersity index and minor distributions of tetrahedral particles. Citrate-capped AgNPs were stable and did not agglomerate upon centrifugation. The effect of AgNPs on biofilm reduction was evaluated using static culture on 96 well microtiter plates. Results showed that AgNPs with the smallest average diameter were the most effective in the reduction of P. aeruginosa biofilm colonies, which accounted for 90% of removal. The biofilm removal activities of the nanoparticles were found to be concentration independent particularly for the concentration within the range of 80 to 200 µg/ml. 

A novel approach to concurrently deliver oral and inhaled drugs as a single formulation is presented. A triple therapy containing theophylline (THEO; orally delivered) with budesonide (BUD) and terbutaline (TERB) (as single and co-spray dried inhaled powders) were prepared as an ordered mix, with THEO acting as a carrier. The aerosolisation performance of THEO formulations containing BUD and TERB alone, physical mix and co-spray dried powder were evaluated using a next generation impactor (NGI). Physico-chemical properties were investigated using electron microscopy, laser diffraction, dynamic vapour sorption and thermal analysis. NGI analysis indicated that >99% of the THEO powder was greater than 4.46 mm, with >90% dissolved within 5 minutes. Particle size analysis showed samples suitable size for inhalation. Thermal and moisture analysis suggested powders to be stable at room temperature up to 70% RH. Aerosol studies indicated different performance of BUD and TERB depending on the mixing procedure. The co-spray dried formulation showed the highest performance, with a fine particle fraction (≤4.46 microns) of BUD and TERB of 34.39 ± 3.56 % and 33.61 ± 5.67 %, respectively. Such observations suggest this multicomponent drug delivery system could be developed to concomitantly deliver oral and inhaled drugs, an approach that, to date does not exist. Ultimately, this technology potentially reduces the requirement for multiple therapies and increases patient compliance.

Purpose: The aim of this study was to characterize the permeability kinetics of salbutamol sulphate, a commonly used 2-agonist in the treatment of asthma exacerbation, across Calu-3 respiratory epithelial cell monolayers in the presence of non-steroidal anti-inflammatory drugs (NSAIDs), as they have been implicated to be able to modulate organic cation transporters (OCT). Methods: Calu-3 cell monolayers were grown in a liquid covered culture (LCC) configuration on 0.33 cm2 Transwell polyester cell culture supports. Monolayers, cultured between 11 and 14 days were evaluated for epithelial resistance, tight junction integrity and expression of OCT using Western blot analysis. The transport of salbutamol across the monolayer was studied as a function of concentration.  Directional transport was investigated by assessing apical-basal (a-b) and basal- apical (b-a) directions. The influence of a non-specific OCT inhibitor (tetraethylammonium, TEA) and three NSAIDs (aspirin, ibuprofen and indomethacin) on the uptake of salbutamol was studied. Results: The flux of salbutamol sulphate increased with increasing concentration, before reaching a plateau suggesting the involvement of a transport mediated uptake mechanism. Western blot analysis detected the presence of OCT1-3 and N1 and N2 sub-types suggesting the presence of functioning transporters. The apparent permeability (Papp) of 0.1 mM salbutamol across the epithelial monolayer displayed directional transport in the a-b direction which was inhibited by  ~70% in the presence of TEA, suggesting OCT mediated uptake. Likewise, the uptake of 0.1 mM salbutamol was decreased in the presence of all three NSAIDs supporting a mechanism whereby NSAIDs inhibit absorption of salbutamol across the bronchial epithelium via effects on the OCT transporters.

Background: Chronic obstructive pulmonary disease (COPD) is characterised by mucus hyper-production. This pathology, together with other inflammatory contributions, leads to airway obstruction and breathing complications. Newer therapeutic approaches are of increased interest, including the use of HMG-CoA reductase inhibitors. Retrospective studies have shown that statins are effective in reducing patient mortality, and blood cytokines levels. These findings suggest statins may also provide a new therapeutic approach in COPD treatment. Purpose: The aim of the present work was to study the transport of SV across Calu-3 epithelial cells and to investigate its pharmacological action with respect to reduction in mucus production. Methods: Calu-3 cells were grown under liquid covered culture (LCC) conditions for transport studies in order to demonstrate the ability of SV to transport across the monolayer. For mucus detection, cells were grown under air interface culture (AIC) conditions. Samples collected for microscope analysis, were stained with alcian blue; images of the stained cell surface were acquired and the mucus was quantified as the RGBB ratio. Results: SV was transported through the cell monolayer and ‘retained’ inside the Calu-3 cells. Colour analysis of stained Calu-3 monolayers microscope-images, showed that chronic administration of SV for 14 days caused a significant inhibition in mucus production. Conclusion: These findings suggest that local delivery of SV directly to the lungs may provide a promising treatment and potential disease management approach of COPD, with significant effects on mucus reduction.